5-nitro-furfural piperazinealkanoyl hydrazones and method of preparation



United States Patent 3,513,165 S-NITRO-FURFURAL PIPERAZINEALKANOYL HY- DRAZONES AND METHOD OF PREPARATION Elena Massarani, Dante Nardi, and Ludwig Degen, Milan,

Italy, assignors to Societe dExploitations Chimiques et Pharmaceutiques Seceph S.A., Lugano, Switzerland No Drawing. Filed Aug. 29, 1968, Ser. No. 756,290 Claims priority, application Switzerland, Sept. 11, 1967, 12,647/ 67 Int. Cl. C07d 51/70 US. Cl. 260-240 3 Claims ABSTRACT OF THE DISCLOSURE Novel 5 nitro furfural piperazinealkanoylhydrazones having antibacterial activity are prepared by condensing hydrazones with S-nitro-furfural and a carboxylic acid of the formula or by reacting an alcohol with a compound of the formula The object of the present invention is new S-nitrofurfural piperazinealkanoylhydrazones of the formula:

in which R is hydrogen or an alkyl or alkenyl radical having 1 to 12 carbon atoms which is unsubstituted or substituted by at least one hydroxyl or aryl group, an aryl radical, an acyl radical or a dialkyl carbamoyl radical, and X is an alkylene radical having one or two carbon atoms, as well as their acid addition salts.

These compounds have an antibacterial activity on gram-positive bacteria, gram-negative bacteria and the mycobacterium tuberculosis.

The compounds in which R is in an alkyl radical having 1 to 4 carbon atoms, a phenyl radical, a benzyl radical, an acetyl radical or an N-diethyl carbamoyl radical and X is a methylene radical, ethylene or ethylidene radical, as well as their acid addition salts, are particularly active.

In accordance with the invention, the compounds of Formula I are prepared by condensing hydrazine, on the one hand, with S-nitro-furfural and, on the other hand, with a carboxylic acid of the formula:

or a functional derivative thereof, preferably a lower alkyl ester such as the ethyl ester.

In one particular embodiment of this process, the ethyl ester of the carboxylic acid of Formula II is first of all condensed with hydrazine hydrate in a slight excess in the presence of ethanol as solvent. Thereupon the hydrazide thus obtained is condsensed with an equimolecular quantity of S-nitro-furfural dissolved in acetic acid. The desired product is thus obtained in the form of the diacetate from which the base can be liberated by treatment with a basic agent such as sodium carbonate.

The ethyl ester of the carboxylic acid of Formula II can be prepared by reaction of a compound of formula ClXCOOC H with a piperazine of formula:

The ethyl esters of the acids of Formula II, with the exception of those in which R is hydrogen or an alkanoyl or dialkyl carbamoyl radical, are also prepared by reacting a sulfonic ester of the alcohol of formula ROH with a compound of formula:

H-N \NXCOOC2H5 (1v As the sulfonic acid ester, p-toluene sulfonic acid ester is advantageously employed. The two starting materials are preferably employed in equimolecular quantities and reacted in the presence of an alkaline agent such as sodium or potassium carbonate in solution in an alcohol of formula ROH. The reaction time is of the order of 20 hours at C.

In accordance with the invention, the same reaction is used to introduce the radical R into a compound of formula:

for obtaining the S-nitro-furfural piperazinealkanoyl hydrazones of Formula I, with the exception of those in which R is hydrogen or an alkanoyl radical or dialkyl carbamoyl radical.

EXAMPLE 1 (l 4methyl-piperazineacetohydrazide A mixture of 186 g. (1 mol) of ethyl 4-methyl-piperazineacetate, 52.5 g. (1.05 mol) of hydrazine hydrate and 200 cc. of ethyl alcohol is heated under reflux for 12 hours.

Thereupon, the solvent is evaporated under reduced pressure to constant weight. There is obtained a solid white residue which can be used directly for the subsequent condensation with the S-nirto-furfural. The yield is a theoretical yield. The product can be purified by distilling it under vacuum and collecting the fraction of 12S C./0.2 mm. Hg (yield 75%). This substance crystallizes in ligroin with a melting points of 87-89 C. It is soluble in water, methanol, ethanol, chloroform and benzene and practically insoluble in ethyl ether. For C7H1SN4O (mol. wt. 172.23).Calculated (percent): C, 48.81; H, 9.36; N, 32.53. Found (percent): C, 49.06; H, 9.48; N, 32.50.

There remains in the distillation flask a tailing crystallizable in ethyl acetate (M.P. 113 C.), identified as being the bis-1,2-(4'-methyl-piperazineacetyl)-hydrazine of formula:

H3ON\ N- C HzCONH-NH C O CHz-N N-C H3 This substance is soluble in water, methanol, ethanol,

and chloroform and is slightly soluble in ethyl ether and benzene. For C H N O (mol wt. 312,415 ).Calculated (percent): C, 53.82; H, 9.03; N, 26.90. Found (percent): C, 53.33; H, 9.16; N, 27.09.

As this by-product is formed primarily during the distillation, it is preferable to use the crude reaction product for the subsequent condensation.

The initial ethyl-4-methyl-piperazineacetate can be prepared in the following manner:

0.1 mol of N-methyl-piperazine dihydrochloride is dissolved in 16 cc. of water, whereupon 50 cc. of acetone or of ethanol and 27.72 g. (0.33 mol.) of sodium bicar- 4 carbonate to the aqueous extract. The precipitate is collected by filtration, dried in air and crystallized in ethyl acetate, leaving the soluble inorganic part. Yield: 10% of the theoretical yield. The dihydrochloride is obtained by acidification of the mother liquors of the crystallization bonate are added. The mixture is stirred for hour, by means of ethanolic HCl. Yield: of the theoretical whereupon 0.1 mol of ethyl chloroacetate is added to it yield. drop by drop. A slightly exothermal reaction is observed. EXAMPLE 2 Heating is effected under reflux for two hours, whereupon filtration of the hot material is effected to eliminate the 10 5 mtro furfural4 methy P1P erflzmeafetohydrazon inorganic salt, and the solvent is then evaporated under 172 11101) of 4'methY1'PlPefallneacetohydl'azlde vacuum and the residue distillei (prepared in the manner described in Example 1) are dissolved in 200 cc. of hot acetic acid, whereupon the (2) 5-n1trofurfural 4-methyl-p1perazineacethydrazone solution is cooled to and there are added, with 172 g. (1 mol) of 4-rnethyl-piperazinoacethydrazide agitation, 141 g. (1 mol) of S-nitro-furfural dissolved in are dissolved in 200 cc. of acetic acid and cooled to 100 cc. of acetic acid. The mixture is agitated at 40 C. C. With agitation, 141 g. (1 mol) of 5-nitro-furfural for 1 /2 hours and then poured into 1000 cc. of ethyl dissolved in 100 cc. of acetic acid, are added. The mixture acetate. is stirred at 40 C. for 1 /2 hours and then poured into The solution is extracted with 4 successive portions of 3000 cc. of ethyl acetate; 10 g. of bleaching carbon are 20 water of 1000, 500, 250 and 150 cc. It can be verified by added, followed by filtration. Upon cooling, a yellow prodchromatography that the ethyl acetate no longer contains uct crystallizes out. It is set aside for 24 hours in an iceany desired hydrazone. The aqueous extracts are combined box and filtered. There are obtained 375 g. of a product and made alkaline with 2000 cc. of a 20% sodium carmelting at 90 C. and consisting of the desired hydrazone bonate solution. The desired hydrazone separates out in dia etate 25 the form of crystalline base and after the solution has been The above diacetate is suspended in 4500 cc. of ethyl set aside for one hour at 20 C., it is collected by filtraacetate and the mixture heated under reflux with agitation tion. The base thus obtained is dried and then crystallized for one hour. 10 g. of carbon are added to the solution in 3000 cc. of ethyl acetate, thus obtaining 250 g. (85% which is then filtered. It is allowed to cool slowly while of the theoretical yield) of the desired product in base agitating and the desired hydrazone monoacetate crysform. M.P. 167-168 C. This substances is yellow, solutallizes out in an amount of 285 g. (yield 80% referred ble in methanol, ethanol, benzene, chloroform and aceto the 5-nitro-furfural). M.P. 123-125" C. This substance tone, slightly soluble in water and practically insoluble in is yellow and soluble in water, methanol and ethanol ethyl ether. For C H N O .Calculated (percent): C, and practically insoluble in ethyl ether. For 48.80; H, 5.80; N, 23.72. Found (percent): C, 48.72;

C12H17N5O4 CH3COOH H, 5.59; N, 23.74. Calculated (percent): 0, 47.32; H, 5.96; N, 19.71. Ei in H O 577 191 111 011,011 360%652 Found (percent): C, 47.57; H, 6.24; N, 19.85. g at 250 3172501116328 This product can be dried for 4 hours at 40 C. in By dissolving the above base in ethyl alcohol and an oven. At higher temperatures or when it is left in air 40 acidulating the solution by means of ethanolic HCl, the for a long time, it loses acetic acid, while its melting dihydrochloride of the above hydrazone is obtained in point at first drops and finally rises to l67l68 C, theoretical yield. M.P. 252 C. dec. This dihydrochloride which corresponds to a total transformation of the hydrais a yellow substance which is soluble in water and praczone into free base. The monoacetate can, however, be tically inso uble in 10 ethyl ether, benzene a kept in a stoppered bottle indefinitely at 20-25 C. ethyl acetate. For C H N O -2HCl.-Calculated (per- In the mother liquors of the crystallization, one can cent): C, 39.14; H, 5.20; N, 19.46; Cl., 19.26. Found recover an additional amount of the desired product by (P isolating it in the form of base or dihydrochloride. The In Table I there are set forth the properties of other base is obtained by extracting the mother liquors (ethyl 5-nitro-furfural piperazinealkanoyl hydrazones of Foracetate) with a small amount of water and precipitating 0 mula I which can be prepared in similar manner. The the base by addition of a saturated solution of sodium symbols R and X have the meaning previously indicated.

TABLE I Calculated Found R- -X M.P. Formula C H N 01 c H N 31 CH CH -CH 7 C H N O .CH CODE 48. oni-oniom- -oHi 99-100 CilHilNZOiJCfiaCOOH 49% IIIIII CHg-CH -CH 52.00 6.55 21.66 .s 2- 237-9 40. 58 6.08 16.90 17.11 73 62 17. 09

CHg-CHz-CHz-CHz- CH2 153-9 H No 21101 53.40 6.87 20.76 53.53 7.30 20.60 225-7 23 5 43.89 0.14 17.00 7.2 CH3-(CH2)11 --CHg- 144-5 CzaHs9N504 61.44 8.74 15.58 .101-

20 -1 58.21 5.70 18.86 .3 @0132- CHz-- 185-7 icwHzlNfiotzHol-mzo {45. 02 5. 07 14. 92 14.77 0? 14.00

N-C- C'Hz -2 CmOggNaO 50.52 6.36 22.09 60.63 6.51 21.89

TABLE III R-N N-X--C o C2H5 Calculated Found R X M.P. B.P. Formula C H N 01 C H N 01 CH3CH2- CH2 137/22n1m.Hg' CH2oN2Oz 59.97 10.07 13.99 59.78 9.84 14.23 011.011.0112- -CHz- 173-9 136-7l18 mm.Hg C11H22N2O2.2HC1 5166. 11 3 13.09 7 9.66 24.50

cua-cn- -CH; 2034 137/15 mm. Hg CliHnNzOz-ZHCI 45.99 3.42 9.75 2469 46.06 8.67 9.84 24.52

CH -(CH2)3- OHz 197-9 144-5 12mm.r1 0.211241920521101 40 55.46 on3 cH.)1 CHz 160-1 0.2 mm. Hg-.. 045111519201 70150 11I34 3123 11177 3140 GH-CH; CHz- 215-17 147/0.3Inm. C11H24N2O2.2HC1 60.45 8.86 10.33 20 54.74 7.56 8.03 19.98

02N -CHz 122-3 01111151930. 57.32 6.53 14 33 57.43 6.55 14.37

011 00- CH4 125/o.1mm.Hg C10H1QN203.HC1 47.90 7.64 11.17 14 14 43.17 7.35 11.33 14.04

N c- OHz-- 1367 138/0.5mm. Hg C13H25N303.HC1 57.54 9.29 15.49 57.65 9.71 15.60 H 50.75 3.51 13.66 11.52 50.54 3.30 13.55 11.45 C111; 0

H, -1 :n- 116 10mm.Hg CmHzuNzOz 59.97 10.07 13.99 60.04 10.35 14.00

The antibacterial activity of some of the S-nitrofurfural piperazinealkanoyl hydrazones obtained by the present invention was determined on the following microorganisms: Escherichia coli 100, Salmonella typhimurium 1090, Proteus vulgaris OX, Micrococcus pyogenes S65 11, Bacillus subtilis ATCC 9466, cultivated in Difco nutrient agar, on Streptococcus pyogenes A88 cultivated in the Difco agar brain heart infusion agar containing an addition of 5% defibrinated guinea pig blood, and on Clostridium novyi cultivated in Difco thioglycolated Mice liquid medium. The results were read after incubation for 18 hours at 35-37 C.

These compounds were also tested against Mycobacterium tuberculosis H37 Re cultivated on the Kirchner-Hermann medium in the presence of 10% beef serum. The readings were effected after 10, 17 and 24 days.

Table IV below gives the minimum active concentrations expressed in gJml. of the compounds tested. The symbols R and X have the meaning previously indicated.

observed within 7 days following the administration of individual doses of the substances in question. The DL values are set forth in Table V.

There were used for the experiments NMRI white mice and Wistar white rats of both sexes. The mice had a body weight of 18-20 g. and the rats 200-250 g.

TABLE IV S. thypi- Proteus M. S. R X E. coli mun'um vulgaris ptlogenes pyoaenea 3111111118 0'. novyz tuberculosis CH -CH 40 40 80 20 2. 5 20 160 CHi- -CH2 CHr- 20 40 10 20 10 160 10 on.- 10 40 10 40 5 s0 30 C1H5- -CHi- 10 10 40 5 10 5 160 40 C3H711- CH2 20 20 80 5 20 5 40 20 (CH3)2CH CH2 10 20 80 5 10 5 160 10 C4Hnn -OH;;- 10 40 80 10 40 5 40 320 C5H5--CHg- CH 40 160 10 2. 5 5 80 20 C5H CH2- 80 80 80 20 40 5 40 40 CH3OO- -OH 2O 80 160 10 5 20 40 (C2H5)2NCO- -CH 80 10 5 10 80 40 Additional tests using S-nitro-furfural-4'-methylpiperazinoacethydrazine were conducted as follows:

(1) Acute toxicity (DL determined on mice and rats The DL was determined by administering the compound to animals of both sexes, which had fasted for about 14 hours.

There were used NMRI white mice and Wistar white rats.

The DL values were calculated by the method of Litchfield and Wilcoxon (1949) on basis of the mortality 7 was administered the same volume of vehicle in place of 9 10 nitro-furfnral-4'-methyl-piperazinoacethydrazone. The (d) Activity of 5 nitro-furfural-4'-methyl-piperazinomortality of the animals within 7 days following the inacethydrazone of the urinary track with Proteus vulgaris fection was obserbed. O.-Infection of the urinary tract was caused by the The results are set forth in Table VI, from which it method described by Hossack (1962) on female rats of a appears that the treatment with 5-nitro-furfural-4-methylbody weight of 200-250 g. piperazinoacethydrazone resulted in a clear protective ac- The infection was effected in the bladder with a small tion, since the animals treated all died at a time later than zinc plate of 3 x 3 x 1 mm. which had been previously the controls. immersed in a culture of Proteus vulgaris O for 24 hours TABLE VL-EXPE RIMENTAL PERITONITIS FROM .Klebsz'ella pneumom'ue Mortality atter No. of Total Compound mice 18 hrs. 20 hrs. 24 hrs. 2 days 3 days mortality Controls 20 9 5 2 4 20 5-nitrofuriural-4-methyl-piperazinoacethydrazone 2O 1 3 3 2 19 (b) Activity of 5 nitro-furfural-4'-methyl-piperazinoat 37 C. in Difco nutrient broth. There was used the acethydrazone on experimental septicemia from Strepto- 20 26/4 strain isolated from urine obtained from subjects coccus haemolyticus 203.-Groups of 10 mice were inaffected by pyelonephritis. The treatment was initiated on fected intraperitoneally with a charge of Streptococcus the fourth day after the infection, the 5-nitro-furfural-4'- haemolyticus which caused the death by sepsis of the methyl-piperazinoacethydrazone being administered by animals within 48 hours from the infection. The treatment stomach tube in a dose of 50 mg./ kg. dissolved in water was carried out by administering the drug in a dose of in an amount of 10 ml./kg. body weight. The treatment 0.25 mM./ kg. by stomach tube immediately after the inwas carried out daily for 36 consecutive days. One group fection and thereupon once a day for 3 consecutive days. of control rats (infected controls) was treated in the As control, there was infected a group of mice which manner described above, but instead of the S-nitro-furwas administered the same volume of vehicle instead of fural-4'-methyl-piperazinoacethydrazone they received an 5 nitro-furfural-4-methyl-piperazinoacethydrazone. The 30 administration of an equal volume of water. To another mortality within 8 days from the infection was noted. group of rats (non-infected controls), the sterile plate 5 nitro furfural 4'-methyl-piperazinoacethydrazone was introduced into the bladder. Fifteen rats were used exerts a statistically very significant protection with refor the group treated with 5-nitro-furfural-4-methylspect to infection from Streptococcus haemolyticus (Table r piperazinoacethydrazone and 14 rats for each control VII). group.

TABLE VII.EXIPERIMENTAL INFECTION WITH Streptococcus haemolylicus 203 Mortality after time in days Dose, No.0f Total Compounds mlVL/kg. mice 1 2 3 8 mortality Controls 10 3 6 1 10 5-nitrofurfural methyl-piperazinoacethydrazone 0.25 10 1 1 2 (0) Activity of 5 nitro-furfural-4'-methyl-piperazino- During theperiod of the experiment, 5 infected control acethydrazone on experimental septicemia from Salmoanimals died (1 on the 2nd day, 3 on the 14th day, and nella typhimurium 1086.--Groups of 10 mice were in- 1 on the 36th day of the treatment). A single treated fected intraperitoneally with a charge of Salmonella typhirat died on the 15th day, while none of the non-infected murium 1086 such as to cause the death of the animals control animals died during the time of the experiment. within 3 days from the infection. The treatment was car- The percentage of the animals which died during the ried out by administering the drug in a dose of 0.25 treatment was 36% in the case of the infected controls, rnM./kg. by stomach tube one hour before and one and while in the case of the animals treated with S-nitroa half hours after the infection and then once a day for furfural 4-methylpiperazinoacethydrazone it was 7% three consecutive days. As control, there was infected 21 (Table IX).

group of mice to which, instead of the 5-nitro-furfural-4- methyl-piperazinoacethydrazone there was administered an e ual amount of vehicle alone. The mortalit of the TABLE IX--EXPERIMENTAL INFEC- treat d animals was noted during the six days zi fter the TION BY gi n i r itiihg SURVIVAL infection and compared with that found for the control Mum animals during the same period of time. d fm-furififi.

The results of the ex eriments are summarize in y- Table VIII, from which it can be noted that S-nitro-furacetligd rz z d rfe fural-4-methyl-piperazinoacethydrazone lengthened the Dead 5 1 time of survival of 50% of the animals (ET 50) in sta- AuveIIIIIIIIIII: 9 14 tistically significant manner as compared with the controls.

TABLE VIII.--EXPERIMENTAL INFECTION BY Salmonella typhimurium 1086, TREATMENT Mortality after- No. of Total Compounds animals 18 hrs. 1 day 2 days 3 days 4 days 5 days 6 days mortality Controls 10 6 2 1 1 10 5-nitr0furiu'ral-4"methyl piperazinoacethydrazone 10 2 6 1 1 1 1 1 2 (3) Therapeutic indications-Infections of the urinary a benzyl radical, an acetyl radical or an N-diethyl-cartract. bamoyl radical.

Dose.50-20O mg. 2-4 times per day by mouth or 3; S-nitro-furfural-4'-methyl-piperazinoacethydrazone.

100 mg. slowly i.v.

What is claimed is: 1. 5-nitro-furfural-piperazinohydrazone of the formula References Cited UNITED STATES PATENTS T l 3,304,303 2/1967 Schmidt et al. 260-247.1 R N N ONH =CH 0 NO, OTHER REFERENCES v 10 Chemical Abstracts, vol. 62, col. 14,672 (1965) (abwhemm: stract of Chen et a1.).

R is hydrogen, an alkyl radical having 1 to 12 carbon wagnepzook Synthetic Organic chemistry, p. 667

atoms, an alkenyl radical having 1 to 12 carbon atoms, John Wiley and Sons (1953) a phenyl radical, a benzyl radical, a phenylethyl radical, a nitrophenyl radical, a hydroxyethyl radical, an 15 JOHN D RANDOLPH Primary Examiner acetyl radical or a lower dialkyl carbamoyl radical,

and X is a methylene, ethylene or ethylidene group.

2. The compound according to claim 1, wherein R is 260-268; 424-250 an alkyl radical of 1 to 4 carbon atoms, a phenyl radical, 20

U.S. Cl. X.R. 

